CN115399294A - Dynamic determination method for liriomyza trifoliata imago CTmax - Google Patents
Dynamic determination method for liriomyza trifoliata imago CTmax Download PDFInfo
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- CN115399294A CN115399294A CN202210873699.7A CN202210873699A CN115399294A CN 115399294 A CN115399294 A CN 115399294A CN 202210873699 A CN202210873699 A CN 202210873699A CN 115399294 A CN115399294 A CN 115399294A
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- 241000594036 Liriomyza Species 0.000 title claims abstract description 63
- 238000000034 method Methods 0.000 title claims abstract description 26
- 230000000630 rising effect Effects 0.000 claims abstract description 13
- 230000000694 effects Effects 0.000 claims abstract description 5
- 241001520143 Liriomyza trifolii Species 0.000 claims description 6
- 238000002224 dissection Methods 0.000 claims description 3
- 239000003550 marker Substances 0.000 claims description 2
- 241001517923 Douglasiidae Species 0.000 claims 4
- 238000005259 measurement Methods 0.000 claims 2
- 208000005392 Spasm Diseases 0.000 claims 1
- 238000011278 co-treatment Methods 0.000 claims 1
- 238000000691 measurement method Methods 0.000 claims 1
- 241000894007 species Species 0.000 abstract description 3
- 241000255925 Diptera Species 0.000 description 10
- 241000594031 Liriomyza sativae Species 0.000 description 8
- 238000002474 experimental method Methods 0.000 description 7
- 241000238631 Hexapoda Species 0.000 description 3
- 239000006185 dispersion Substances 0.000 description 3
- 230000003028 elevating effect Effects 0.000 description 3
- 239000011148 porous material Substances 0.000 description 3
- 230000009286 beneficial effect Effects 0.000 description 2
- 235000013601 eggs Nutrition 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 241000590412 Agromyzidae Species 0.000 description 1
- 206010052428 Wound Diseases 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 239000003337 fertilizer Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 230000001678 irradiating effect Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000000737 periodic effect Effects 0.000 description 1
- 230000001932 seasonal effect Effects 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/033—Rearing or breeding invertebrates; New breeds of invertebrates
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- Life Sciences & Earth Sciences (AREA)
- Environmental Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Zoology (AREA)
- Animal Husbandry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses a dynamic determination method of a liriomyza trifoliata imago CTmax, which comprises the following steps: (1) Using CO 2 Treating adult liriomyza trifoliata and subpackaging the adult liriomyza trifoliata into a porous plate; (2) keeping the temperature of the program lifting thermostat stable; (3) After the adult liriomyza trifoliata resumes normal activity, placing the 24-hole plate in a program lifting thermostat, and setting a temperature rising slope and a highest temperature; (4) Starting a computer for video recording, stopping the procedure of lifting the thermostat and carrying out computer video recording when all the adult liriomyza trifoliata cannot move, and taking out the porous plate; and (5) confirming the CTmax value of the leaf-spot liriomyza imago. The method adopts common laboratory instruments, and is simple to operate; the high-temperature tolerance of the liriomyza trifoliata imagoes at the dynamic temperature can be more accurately measured, and the method has a certain reference value for closely related species of the liriomyza trifoliata imagoes.
Description
Technical Field
The invention belongs to the field of entomology, and particularly relates to a dynamic determination method of a leaf-shaped leaf-spot-fly adult CTmax.
Background
The Liriomyza trifoliata (Liriomyzatrifii) belongs to Diptera of Diptera, agromyzidae, phytomyzidae of Heriomyidae, wherein the larvae are harmed by predatory leaves, the adults are harmed by taking food and laying eggs on the leaves, and meanwhile, the plant wounds caused by the feeding and laying eggs of the adults also provide infection ways for germs. At present, the fertilizer is widely distributed in places such as Hainan, yunnan, fujian, shandong, anhui, jiangsu and the like, and causes great harm to the production of vegetables and flowers in China. In the process of seasonal increase of temperature and gradual increase of temperature in the daytime, whether the heat resistance of the liriomyza trifoliata is changed or not reflects the strength of the high-temperature adaptability of the liriomyza trifoliata, so that the summer distribution of the liriomyza trifoliata population is influenced. At present, most of research methods about the relationship between high temperature and liriomyza trifoliata are used for researching the life history characters of the liriomyza trifoliata, such as development, reproduction, survival and the like, under constant temperature, and the liriomyza trifoliata has periodic changes of day and night and seasons in the temperature of the field environment, so that the biological data obtained by a constant high temperature test is used for explaining the field population dynamics and is easy to deviate.
Critical thermal maximum (CTmax) refers to the upper temperature limit of heat resistance of an insect at which the insect will cramp, lose its ability to move, and eventually die. At present, the index is mainly used for measuring the heat resistance difference between various insect states and different geographical populations.
Disclosure of Invention
The purpose of the invention is as follows: the invention aims to provide a simple method for measuring the CTmax of the adult liriomyza sativae, which can more accurately measure the high-temperature tolerance of the adult liriomyza sativae at a dynamic temperature.
The technical scheme is as follows: the invention relates to a dynamic determination method of liriomyza trifolii CTmax, which comprises the following steps:
(1) Using CO 2 Treating the adult liriomyza trifoliata to make the adult liriomyza trifoliata incapable of moving, and subpackaging the adult liriomyza trifoliata into a porous plate;
(2) Setting the temperature of the program lifting thermostat to be suitable for feeding the adult liriomyza trifoliata, and stably keeping the temperature in the thermostat;
(3) Subjecting to CO as described in step (1) 2 After the treated adult liriomyza trifoliata flies recover to normal activities, the porous plate filled with the adult liriomyza trifoliata flies is placed in the procedure lifting thermostat of the step (2), and the temperature is setDegree rise slope and maximum temperature;
(4) Starting a temperature rising program, starting a computer video recording when the temperature starts to rise, stopping the program lifting thermostat and the computer video recording when all the liriomyza trifoliata adults in the porous plate in the step (3) cannot move, and taking out the porous plate;
(5) And (4) observing the time point when the adult liriomyza trifoliata flies reach the critical state and the corresponding temperature of the time point of the critical state recorded by the program lifting thermostat through the computer video in the step (4), and confirming the highest tolerance temperature of each adult liriomyza trifoliata fly in the porous plate to obtain the CTmax value of the adult liriomyza trifoliata flies.
Preferably, the porous plate is a 12-well plate or a 24-well plate.
Preferably, the multi-well plate is pretreated before the determination in the step (1), specifically, a vent hole is punched by using a cover corresponding to each hole of the multi-well plate through dissection, a light source is used for irradiating the multi-well plate, the hole with the brightest reflection is marked by a marker pen, and no adult liriomyza sativae is placed in the hole.
Preferably, the temperature suitable for breeding the adult liriomyza trifoliata in the step (2) is 25-27 ℃.
Preferably, in the step (3), the standard for judging the normal activity of the leaf spot liriomyza sativae adults is that the perforated plates are turned upside down, and the leaf spot liriomyza sativae adults do not fall off.
Preferably, the temperature rising slope of the step (3) is set to be 0.1 ℃/min to 1 ℃/min.
Further preferably, the temperature rising slope of step (3) is set to 0.5 ℃/min
Preferably, the maximum temperature in step (3) is set to 50 to 60 ℃.
Preferably, the criterion for determining that the liriomyza trifoliata adults in the step (5) reach the critical state is that when the liriomyza trifoliata adults suddenly fall from the hole wall to the bottom, the body of the liriomyza trifoliata adults is spasticized.
Preferably, the diameter of the hole of the 24-hole plate is 16mm, and the depth of the hole is 17mm.
Has the advantages that: compared with the prior art, the invention has the following remarkable advantages: (1) The CTmax of the liriomyza trifoliata imago can be quickly and effectively determined, and the CTmax determination of the liriomyza trifoliata imago, south America liriomyza sativae imago also has certain reference value for the closely related species of the liriomyza trifoliata, south America liriomyza sativae imago; (2) A 24-hole plate, a computer and a program lifting thermostat are adopted, are common instruments in a laboratory and are simple to operate; (3) The high-temperature tolerance capability of the liriomyza sativae adults at the dynamic temperature can be more accurately measured by setting different temperature rising slope parameters.
Drawings
FIG. 1 is a schematic diagram of a 24-well plate, wherein A is a 24-well plate body and B is a 24-well plate cover;
FIG. 2 is a schematic view of a program lift incubator;
FIG. 3 is a schematic view of a 24-well plate placed in a process lift oven;
FIG. 4 is a computer observed image;
FIG. 5 is a boxplot of CTmax values for leaf-spot-fly adults.
Detailed Description
For a further understanding of the present invention, reference will now be made in detail to the embodiments illustrated in the drawings.
Example 1
(1) As shown in fig. 1 and 5, 2 air holes are punched by using a cover corresponding to each hole of a 24-hole plate through dissection, the 24-hole plate is irradiated by a light source, the second row and the third row of holes with the brightest reflection are marked by a marking pen, and in order to avoid inaccurate observation, no adult liriomyza trifoliata flies are placed in the holes.
(2) Using CO 2 And (3) treating 5 liriomyza trifoliata adults to enable the liriomyza trifoliata adults to be incapable of moving, and subpackaging the liriomyza trifoliata adults into 24-hole plates, wherein each hole is provided with a diameter of 16mm and a depth of 17mm.
(3) The programmed elevator oven temperature as shown in fig. 2 was set to 25 deg.c and the temperature was stably maintained in the oven.
(4) Turning the 24-hole plate upside down to prevent the leaf-spot liriomyza imagoes from falling, namely, when the leaf-spot liriomyza imagoes recover to normal movement, as shown in figure 3, placing the 24-hole plate containing the leaf-spot liriomyza imagoes into a program lifting thermostat, and setting the temperature rising slope to be 0.1 ℃/min and the highest temperature to be 55 ℃;
(5) Starting the computer video recording shown in the figure 4 when the temperature starts to rise, stopping the procedure of lifting the thermostat and carrying out the computer video recording when 5 liriomyza trifoliata adults in the 24 pore plates all fall to the bottom from the pore wall and the polypide is spasticized, and taking out the 24 pore plates;
(6) And observing the time point when the adult liriomyza trifoliata flies reach the critical state and the corresponding temperature of the time point of the critical state recorded by the program lifting thermostat through a computer video, and confirming the highest tolerance temperature of each adult liriomyza trifoliata flies in the porous plate to obtain the CTmax value of the adult liriomyza trifoliata flies.
The experiment was repeated three times and the results are shown in figure 5.
Example 2
Similar to the procedure of example 1, except that the temperature rising slope of the programmed elevating incubator was set to 0.5 ℃/min. Finally obtaining the CTmax value of the leaf spot liriomyza imago.
The experiment was repeated three times, and the results are shown in FIG. 5.
Example 3
Similar to the procedure of example 1, except that the temperature rising slope of the programmed elevating incubator was set to 1 ℃/min. Finally obtaining the CTmax value of the leaf spot liriomyza imago.
The experiment was repeated three times and the results are shown in figure 5.
Example 4
Similar to the procedure of example 1, except that the programmed elevating thermostat was set to have an initial temperature of 26 ℃ and an end temperature of 50 ℃; finally obtaining the CTmax value of the leaf spot liriomyza imago.
The experiment was repeated three times, and the results are shown in FIG. 5.
Example 5
The procedure of example 1 was followed except that the programmed temperature raising and lowering incubator was set to 27 ℃ for the initial temperature and 60 ℃ for the final temperature. Finally, the CTmax value of the leaf spot liriomyza imago is 5.
The experiment was repeated three times, and the results are shown in FIG. 5.
As can be seen from FIG. 5, the dispersion degree of the treatment values of 0.1 ℃/min and 1 ℃/min is larger than that of 0.5 ℃/min, and the treatment time of 0.1 ℃/min is about 4 hours, so that the method has the characteristic of poor repeatability. While the treatment at 1 deg.C/min requires about 20 minutes, the data dispersion degree is too large. Whereas a treatment of 0.5 c/min requires about 40 minutes and the degree of dispersion of the data is small.
Research results show that newly emerged liriomyza trifoliata adults have relatively strong vitality and can better embody the temperature tolerance of one population. In this experiment, it was found that: the temperature rising slope of 1 ℃/min causes large difference between experimental data and lacks of relevance; the temperature rising slope of 0.1 ℃/min leads to the rapid increase of the experimental time, which is not beneficial to the large-batch experimental treatment; the temperature rising slope of 0.5 ℃/min, on one hand, the measured data accords with the actual situation of species tolerance, and on the other hand, the used time is shorter, which is beneficial to repeat in large batch. In summary, the temperature rise slope of 0.5 ℃/min is the optimal temperature rise slope in this experiment.
Claims (8)
1. A dynamic determination method for a Liriomyza trifolii imago CTmax is characterized by comprising the following steps:
(1) Using CO 2 Treating the adult liriomyza trifoliata to make the adult liriomyza trifoliata incapable of moving, and subpackaging the adult liriomyza trifoliata into a porous plate;
(2) Setting the temperature of the program lifting thermostat to be suitable for feeding the adult liriomyza trifolii, and stably keeping the temperature in the thermostat;
(3) Subjecting to CO treatment in step (1) 2 After the treated adult liriomyza trifoliata recovers normal activity, placing the porous plate filled with the adult liriomyza trifoliata into the program lifting thermostat of the step (2), and setting a temperature rising slope and the highest temperature;
(4) Starting a temperature rising program, starting a computer video recording when the temperature starts to rise, stopping the program lifting thermostat and the computer video recording when all the liriomyza trifoliata adults in the porous plate in the step (3) cannot move, and taking out the porous plate;
(5) And (5) observing the time point when the Liriomyza trifoliata imagoes reach the critical state and the corresponding temperature of the time point of the critical state recorded by the program lifting thermostat through the computer video in the step (4), and confirming the highest tolerance temperature of each Liriomyza trifoliata imagoes in the porous plate to obtain the CTmax value of the Liriomyza trifoliata imagoes.
2. The method for dynamically measuring the CTmax of the adult liriomyza trifoliata, wherein the multi-well plate is a 12-well plate or a 24-well plate.
3. The dynamic measurement method for the leaf-spot fly imago CTmax as claimed in claim 1, characterized in that the multi-well plate is pretreated before the measurement in step (1), specifically, a vent hole is pricked by a cover corresponding to each well of the multi-well plate through dissection, the multi-well plate is irradiated by a light source, the well with the brightest light reflection is marked by a marker pen, and the leaf-spot fly imago is not placed in the well.
4. The method for dynamically measuring the adult liriomyza trifolii CTmax as claimed in claim 1, wherein the temperature suitable for feeding the adult liriomyza trifolii CTmax in step (2) is 25-27 ℃.
5. The method for dynamically measuring the leaf-miners CTmax as claimed in claim 1, wherein in the step (3), the criterion for judging the return of the leaf-miners to normal activity is that the perforated plate is turned upside down and the leaf-miners do not fall.
6. The method for dynamically measuring the adult CTmax of liriomyza trifoliata as claimed in claim 1, wherein the temperature rise slope in step (3) is set to 0.1 ℃/min to 1 ℃/min.
7. The method for dynamically measuring the adult leaf miner CTmax as claimed in claim 1, wherein the maximum temperature in the step (3) is set to 50-60 ℃.
8. The method for determining the dynamic measurement of the liriomyza trifoliata imago CTmax as claimed in claim 1, wherein the criterion of the criterium trifoliata imago reaching the critical state in the step (5) is that when the liriomyza trifoliata imago suddenly falls from the hole wall to the bottom, the body of the liriomyza trifoliata imago is in spasm.
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